In electrospray ionization, a liquid is sprayed through the tip of a needle that is held at a high electric potential of a few kilovolts. Small multiply-charged droplets containing solvent molecules and analyte molecules are initially formed and then shrink as the solvent molecules evaporate. The shrinking droplets also undergo fission—possibly multiple times—when the shrinkage causes the charge density of the droplet to increase beyond a certain threshold. This process ends when all that is left of the droplet is a charged analyte ion that can be mass analyzed by a mass spectrometer. Some of the droplets and liberated ions are directed into the vacuum chamber of the mass spectrometer through an ion inlet orifice, such as an ion transfer tube that is heated to help desolvate remaining droplets or ion/solvent clusters. A strong electric field in the tube lens following the ion transfer tube also aids in breaking up solvent clusters. The smaller the initial size of the droplets, the more efficiently they can be desolvated, and eventually, the more sensitive the mass spectrometer system becomes.
One of the design parameters that influence the initial size of the droplets is the size of the emitter orifice through which they are being formed. So-called nanospray ionization is a form of electrospray ionization that employs small-diameter tips in the order of tens of micrometers. This limits the maximum solvent flow rates to the range of tens of microliters to nanoliters per minute. It is well known in the art that, of all the variants of electrospray ionization, nanospray ionization yields the highest current per analyte concentration. This result is attributed to the small bore of the electrospray emitter needles employed, which cause the diameter of the droplets formed at the Taylor cone to be the smallest, such that the combined effects of smaller initial droplet size and higher analyte concentration (as a result of less required solvent) permit a higher proportion of ions to be inlet into a mass spectrometer. Therefore, nanospray ionization enables the most sensitive results to be obtained from a mass spectrometer.
Unfortunately, due to the small-diameter emitter needles employed in nanospray ionization, there is a maximum to the amount of liquid flow that can be accommodated. Therefore, nanospray is limited in its applications to low flow analysis. However, in LC-MS (Liquid Chromatography-Mass Spectrometry) assays, much larger flow rates are encountered, often exceeding 100 microliters per minute and occasionally as high as 5 milliliters per minute. For those flow rates, larger bore needles are conventionally employed and the electrospray variant with pneumatic assist (“sheath” or nebulizing gas) is used to enable shearing off of droplets from the liquid stream as well as to cause subsequent breakdown of the large droplets. The sheath gas may be heated in order to expedite de-solvation. Often, additional auxiliary gas flows (which could be heated) are employed to help the ions escape from the larger solvent droplets.
FIG. 1 illustrates a conventional electrospray system having pneumatic assist, as taught in U.S. Pat. No. 4,861,988 in the names of Henion et al. The instrument system 1 includes an atmospheric pressure ionization chamber 2, a gas curtain chamber 3 and a vacuum chamber 4. The ionization chamber 2 is separated from the gas curtain chamber 3 by an inlet plate 5 containing an inlet orifice 6. The gas curtain chamber 3 is separated from the vacuum chamber 4 by an orifice plate 7 containing an orifice 8. The gas curtain chamber 3 is supplied from a source 11 with a curtain gas (typically nitrogen or argon) at a pressure higher than that prevailing in the ionization chamber 2. In use, the sample to be analyzed is introduced into the ionization chamber 12 and is ionized. The ions are drawn by an electric field through the inlet opening 6, through the orifice 8, and are focused by a lens 9 into a mass spectrometer 10.
Still referring to FIG. 1, liquid from a small-bore liquid chromatograph 12 flows through a thin quartz tube 13 into an “ion spray” device 14. The ion spray device 14 comprises a stainless steel capillary tube 15 of circular cross-section, encircled by an outer tube 16 also of circular cross-section. The inner diameter of the stainless steel capillary tube 15 is typically 0.1 millimeters, and its outer diameter is typically 0.2 millimeters. The inner diameter of the outer tube 16 is typically 0.25 millimeters, leaving an annular space 31 between the two tubes of thickness 0.025 mm. Normally, the tip of the stainless steel tube 15 protrudes slightly from the outer tube 16.
Typically the quartz tube 13 from the liquid chromatograph 12 will be 0.050 mm inner diameter. The tube 13 is sealed at its end 35 to the stainless steel tube 15, so that the liquid flowing in the tube 13 can expand into the stainless steel tube.
A gas, typically nitrogen boiled from liquid nitrogen, is introduced into the space 31 between the tubes 15, 16 from a gas source 17. The gas source 17 is connected to the outer tube 16 by a fitting 18, through which the inner quartz tube 13 passes. Other gases, such as “zero air” (i.e. air with no moisture) or oxygen can also be used.
A source 19 of electric potential is connected to the stainless steel tube 15. For negative ion operation, the stainless steel capillary may be kept at −3000 volts, and for positive ion operation at +3000 volts. The orifice plate 5 is grounded. In operation of the apparatus 1, charged droplets are emitted from the end of the stainless steel tube 15 by electrospray ionization at the same time that the gas flows through the space 31 surrounding the stainless steel tube 15. The combination of the electric field and the gas flow serves to nebulize the liquid stream. The nebulizer gas flow through the annular space 31 also allows a larger distance to be maintained between the tip of the stainless steel tube 15 and the orifice plate 5 than in the case when no gas is used, thus helping to reduce the electric field at the tip of the tube and prevent corona discharge.
Various designs have been proposed in an attempt to extend the benefits of small initial droplets—as are associated with low flow rates, for example, nanospray—to the larger flow rates required for LC-MS analysis. The concept is to use multiple low-flow rate emitters in parallel so as to divide the large flow into a large number of smaller flows, each directed to a single emitter. An example of an apparatus that employs this strategy is shown in FIG. 2, in which is illustrated an array of fused-silica capillary nano-electrospray ionization emitters arranged in a circular geometry, as taught in United States Patent Application Publication 2009/0230296 A1, in the names of Kelly et al. Each nano-electrospray ionization emitter 21 comprises a fused silica capillary having a tapered tip 22. As taught in United States Patent Application Publication 2009/0230296 A1, the tapered tips can be formed either by traditional pulling techniques or by chemical etching and the radial arrays can be fabricated by passing approximately 6 cm lengths of fused silica capillaries through holes in one or more discs 20. The holes in the disc or discs may be placed at the desired radial distance and inter-emitter spacing and two such discs can be separated to cause the capillaries to run parallel to one another at the tips of the nano-electrospray ionization emitters and the portions leading thereto. Analogous benefits have been described by Smith and coworkers in U.S. Pat. No. 6,831,274 (combination of multiple electrosprayers with an ion funnel).
An issue with having a multitude of nanospray emitters is that the generated cloud of droplets starts to have dimensions that become incompatible with those of the inlet orifice of the mass spectrometer, in other words only a fraction of the mist generated is actually drawn into the inlet of the mass analyzer. This loss obviously results in decreased sensitivity of the instrument. Some possible remedies to this problem could be to provide larger or additional inlets to the mass spectrometer, but that in turn causes a larger (or more) vacuum pump(s) to be required to maintain similar pressures in the mass spectrometer. This leads to additional costs, spatial requirements, shipping weight etc. all of which are not beneficial.
In considering emitter arrays, it is desirable to be able to balance the desirable effects of small low-flow-rate emitters against the possible undesirable effects of a large number of emitters. In order to divide the total flow from a conventional liquid chromatograph among several emitters interfaced to a conventional mass spectrometer ion inlet, the distance between the individual emitters should be maintained as small as possible. However, it is also known in the art that, in order for a Taylor cone to be formed, a high electric field gradient is required. Commonly, this is obtained by having a high aspect ratio structure such as a needle. Yet, when there are multiple needles in close proximity, the spray from one needle could be negatively impacted by the electric field around a neighboring needle. Also, when multiple emitters abut one another, because of the surface tension, the eluent from the different channels could coalesce rather than form individual Taylor cones. All such issues could be resolved by using a limited number of emitters—such that the flow rate per emitter is in the range of hundreds of microliters to a few milliliters per minute—in conjunction with pneumatic assist techniques.
Arrays of electrospray emitters in close proximity to one another are known in the art. Microfabrication techniques that have been borrowed from the electronics industry and microelectromechanical systems (MEMS), such as chemical vapor deposition, molecular beam epitaxy, photolithography, chemical etching, dry etching (reactive ion etching and deep reactive ion etching), molding, laser ablation, etc., have been used to fabricate such emitter arrays. For instance, FIGS. 3A-3B show, respectively, a schematic view of one electrospray system and a cross-sectional view of an electrospray device of the system, as taught in United States Patent Application Publication 2002/0158027 A1, in the names of Moon et al. The individual electrospray device 204, which may comprise one member of an array of such devices, generally comprises a silicon substrate or microchip or wafer 205 defining a channel 206 through substrate 205 between an entrance orifice 207 on an injection surface 208 and a nozzle 209 on an ejection surface 210. The nozzle 209 has an inner and an outer diameter and is defined by a recessed region 211. The region 211 is recessed from the ejection surface 210, extends outwardly from the nozzle 209 and may be annular. The tip of the nozzle 209 does not extend beyond the ejection surface 210 to thereby protect the nozzle 209 from accidental breakage.
A grid-plane region 212 of the ejection surface 210 is exterior to the nozzle 209 and to the recessed region 211 and may provide a surface on which a layer of conductive material 214 including a conductive electrode 215 may be formed for the application of an electric potential to the substrate 205 to modify the electric field pattern between the ejection surface 210, including the nozzle tip 209, and the extracting electrode 217. Alternatively, the conductive electrode may be provided on the injection surface 208 (not shown).
The electrospray device 204 further comprises a layer of silicon dioxide 213 over the surfaces of the substrate 205 through which the electrode 215 is in contact with the substrate 205 either on the ejection surface 210 or on the injection surface 208. The silicon dioxide 213 formed on the walls of the channel 206 electrically isolates a fluid therein from the silicon substrate 205 and thus allows for the independent application and sustenance of different electrical potentials to the fluid in the channel 206 and to the silicon substrate 205. Alternatively, the substrate 205 can be controlled to the same electrical potential as the fluid.
As shown in FIG. 3A, to generate an electrospray, fluid may be delivered to the entrance orifice 207 of the electrospray device 204 by, for example, a capillary 216 or micropipette. The fluid is subjected to a electrical potential Vfluid via a wire (not shown) positioned in the capillary 216 or in the channel 206 or via an electrode (not shown) provided on the injection surface 208 and isolated from the surrounding surface region and the substrate 205. An electrical potential Vsubstrate may also be applied to the electrode 204 on the grid-plane 212, the magnitude of which is preferably adjustable for optimization of the electrospray characteristics. The fluid flows through the channel 206 and exits or is ejected from the nozzle 209 in the form of very fine, highly charged fluidic droplets 218. The extracting electrode 217 may be held at an electrical potential Vextract such that the electrospray is drawn toward the extracting electrode 217 under the influence of an electric field.
Almost all microfabricated electrospray nozzles or other emitters have no provision for delivery of a nebulizing gas directly to the nozzle or emitter. One apparatus that is an exception to this statement is disclosed in United States Patent Application Publication 2006/0113463 A1 in the names of Rossier et al., as is illustrated in FIG. 4. The apparatus 23 illustrated in FIG. 4 is made in a substrate 24 and comprises two covered microstructures, namely a sample microchannel 25 and a sheath liquid microchannel 26 that are connected to inlet reservoirs 27, 28 respectively, placed on the same side of the support 24 for fluid introduction. The microstructures have an outlet 29 formed at the edge of the support, at which the spray is to be generated upon voltage application.
As described in the aforementioned United States Patent Application Publication 2006/0113463 A1, the apparatus 23 comprises two plasma etched microchips made of a polyimide foil having a thickness of 75 μm, comprising one microchannel (approximately 60 μm×120 μm×1 cm) sealed by lamination of a 38 μm thick polyethylene/polyethylene terephthalate layer and one gold microelectrode (not illustrated) of approximately 52 μm diameter integrated at the bottom of the microchannel. The two polyimide chips are glued together and further mechanically cut in a tip shape, in such a manner that this multilayer system exhibits two microstructures both comprising a microchannel having an outlet at the edge of the polyimide layers, thereby forming an apparatus such that the outlets of the sample and sheath liquid microstructures are superposed. The thickness of the support separating the two microstructure outlets may be less than 50 micrometers.
In operation of the apparatus 23, when an electrical potential is applied to the electrode, a Taylor cone is formed that encompasses the outlets 29 of both the sample and sheath liquid microchannels, so that the sample solution mixes with the sheath liquid solution directly in the Taylor cone. Rossier et al. further teach that, instead of a sheath liquid, a sheath gas may be introduced into the micro-channel 26. This gas may be an inert gas such as nitrogen, argon, helium or the like, serving e.g. to favor the spray generation and/or to prevent the formation of droplets at the microstructure outlet. For some applications, a reactive gas such as oxygen or a mixture of inert and reactive gases may also be used so as to generate a reaction with the sample solution. Rossier et al. further teach that an array of such apparatuses can be constructed.
Likewise, United States Patent Application Publication US 2007/0257190 A1, in the name of inventor Li, teaches microfluidic chip structures for gas assisted ionization, these structures having an analyte channel ending in a spray tip and having up to four gas channels having outlet ends adjacent to the spray tip. For instance, Li teaches an apparatus having a spray tip having a first pair of gas channels having ends disposed at opposite sides of the spray tip and a second pair of gas channels, provided by auxiliary gas chips, also disposed at opposite ends of the spray tip.
Although the apparatuses taught by Rossier et al. and by Li appear to operate adequately, they only provide for introduction of a sheath gas at a finite number of discrete gas channel ends adjacent to a fluid channel. The nebulizing gas provided by these finite numbers of discrete gas channels thus does not exit the channels in a fashion that two-dimensionally circumferentially surrounds the fluid emitted from the fluid channel. As a result, these apparatuses are subject to potential asymmetry or non-uniformity in the sheath pressure or flow rate around the emitted droplets or other charged particles. For instance, if the sheath or nebulizing gas is supplied via a single channel aperture on one side of the Taylor cone, the supplied gas flow may not symmetrically surround the stream of emitted droplets. If the gas is supplied from multiple channels, then restricted flow or clogging in one or more of the channels may cause similar difficulties. Since sheath gas is supplied under pressure, the introduction of sheath gas in such an asymmetric or non-uniform fashion in such existing apparatuses, if not carefully controlled, may perturb the emission pattern and direction of electrospray droplets in a manner that causes fluctuations in the ability of ions to be captured by an ion inlet port of a mass spectrometer. Further, since the outlets of both the sample and sheath liquid or gas microchannels, as described in the Rossier et al. apparatus, must fit within the dimensions of an individual Taylor cone, this apparatus is limited to nanospray flow regimes and is not suitable for providing variable flow rates in the range of hundreds of microliters to a few milliliters per minute, as would be expected when dividing a total sample flow of an LC-MS among a limited number of emitters.